Analysis of proenkephalin A, proopiomelanocortin and protachykinin neuropeptides in human lumbar cerebrospinal fluid by reversed-phase high-performance liquid chromatography, radioimmunoassay and enzymolysis
Harajiri, S.; Wood, G.; Desiderio, D.M.
Journal of Chromatography 575(2): 213-222
1992
ISSN/ISBN: 0021-9673 PMID: 1629297 Document Number: 398200
A comprehensive high-performance liquid chromatographic, radioimmunoassay, and enzymatic degradation scheme has been developed to analyze several intact neuropeptides and and the corresponding peptides created by in vivo enzymolysis of precursors to study neuropeptides in human lumbar cerebrospinal fluid (CSF) and to test the hypothesis that defects in the metabolism (synthesis, degradation) of neuropeptide precursors, neuropeptides, and metabolites play a role in low back pain. CSF samples were obtained from three different patient groups: controls (C), whose low back pain was relieved without lidocaine; pharmacological responders (PR), whose pain was relieved by lidocaine and who were candidates for surgery; and pharmacological non-responders (PNR), whose pain was not relieved by lidocaine and a mid-thoracic anesthetic, and who were not candidates for surgery. The metabolic activity involved during synthesis and degradation of the peptides was assessed by measuring intact, native neuropeptide immunoreactivity in pre-incubated and post-incubated CSF samples, where samples were incubated at 37.degree. C for 1 h. Pre-incubation radioimmunoassay measurements reflected the content of intact peptides present in lumbar CSF at the time of sampling, and post-incubation measurements assayed the amount of peptide that had remained embedded within its precursors [cryptic methionine enkephlin (ME)] and that had been released by the action of CSF peptidases. Significant differences were found in post-incubation samples for the amount of proenkephlin A [ME, leucine enkephalin (LE)] and tachykinin [substance P (SP)] peptides. For example, significant differences were observed for ME-like immunoreactivity (C versus cryptic), SP-like immunoreactivity (PNR versus PR), and LE-like immunoreactivity (PR versus C). No significant differences were observed among the peptides within the pre-incubation samples. The significant differences observed within the post-incubation samples demonstrated that the neuropeptides that had remained encrypted within their precursors in the corresponding pre-incubated samples may reflect an altered metabolism of the proenkephalin A (ME, LE) and tachykinin (SP) systems, and therefor those neuropeptide families may be significant factors in idiopathic low back pain.